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1.
Front Immunol ; 14: 1252765, 2023.
Article in English | MEDLINE | ID: mdl-37809070

ABSTRACT

Background: Bruton's tyrosine kinase (BTK) is a cytoplasmic protein involved in the B cell development. X-linked agammaglobulinemia (XLA) is caused by mutation in the BTK gene, which results in very low or absent B cells. Affected males have markedly reduced immunoglobulin levels, which render them susceptible to recurrent and severe bacterial infections. Methods: Patients suspected with X-linked agammaglobulinemia were enrolled during the period of 2010-2018. Clinical summary, and immunological profiles of these patients were recorded. Peripheral blood samples were collected for monocyte BTK protein expression detection and BTK genetic analysis. The medical records between January 2020 and June 2023 were reviewed to investigate COVID-19 in XLA. Results: Twenty-two patients (from 16 unrelated families) were molecularly diagnosed as XLA. Genetic testing revealed fifteen distinct mutations, including four splicing mutations, four missense mutations, three nonsense mutations, three short deletions, and one large indel mutation. These mutations scattered throughout the BTK gene and mostly affected the kinase domain. All mutations including five novel mutations were predicted to be pathogenic or deleterious by in silico prediction tools. Genetic testing confirmed that eleven mothers and seven sisters were carriers for the disease, while three mutations were de novo. Flow cytometric analysis showed that thirteen patients had minimal BTK expression (0-15%) while eight patients had reduced BTK expression (16-64%). One patient was not tested for monocyte BTK expression due to insufficient sample. Pneumonia (n=13) was the most common manifestation, while Pseudomonas aeruginosa was the most frequently isolated pathogen from the patients (n=4). Mild or asymptomatic COVID-19 was reported in four patients. Conclusion: This report provides the first overview of demographic, clinical, immunological and genetic data of XLA in Malaysia. The combination of flow cytometric assessment and BTK genetic analysis provides a definitive diagnosis for XLA patients, especially with atypical clinical presentation. In addition, it may also allow carrier detection and assist in genetic counselling and prenatal diagnosis.


Subject(s)
Agammaglobulinemia , COVID-19 , Male , Pregnancy , Female , Humans , Protein-Tyrosine Kinases/genetics , Malaysia , COVID-19/genetics , Agammaglobulinaemia Tyrosine Kinase/genetics , Agammaglobulinemia/diagnosis , Agammaglobulinemia/genetics
2.
Front Immunol ; 14: 1209315, 2023.
Article in English | MEDLINE | ID: mdl-37529038

ABSTRACT

Introduction: With increased diagnostic capabilities and treatment modalities in the field of primary immunodeficiencies (PID), many pediatric patients survive beyond childhood and experience a change of care to the adult-oriented healthcare system. Unfortunately, the transition pathways for PID are less clearly defined, resulting in deterioration of quality of care in adulthood. Hence, this is the first regional study to address PID clinicians' opinions on practices and challenges of transition care in 7 Southeast Asia (SEA) countries. Methods: We adopted a cross-sectional study design through an online survey platform to enquire opinions of transition practices from expert representatives in 7 SEA countries. Results: Regionally, 3 out 7 countries reported having no practice of transition care. Among cited challenges were reluctant adaptation by patients and caregivers to unfamiliarized adult healthcare systems, inadequate ratio of adult immunologists to patients and lack of facilities for transfer. Discussion and conclusion: Our study provides evidence to advocate policy makers on the importance of standardized integration of transition practice towards betterment of transiting PID patients into adulthood.


Subject(s)
Primary Immunodeficiency Diseases , Adult , Child , Humans , Asia, Southeastern/epidemiology , Cross-Sectional Studies , Primary Immunodeficiency Diseases/diagnosis , Primary Immunodeficiency Diseases/epidemiology , Primary Immunodeficiency Diseases/therapy , Surveys and Questionnaires , Transition to Adult Care
3.
JMIR Res Protoc ; 12: e46816, 2023 Jun 26.
Article in English | MEDLINE | ID: mdl-37358892

ABSTRACT

BACKGROUND: Allergic diseases affect around 40% of the pediatric population worldwide. The coexistence of asthma, allergic rhinitis, eczema, and food allergy renders allergy treatment and prevention challenging. Infant feeding strategies recommend avoiding allergenic foods to prevent allergy development and anaphylaxis. However, recent evidence suggests that early consumption of food allergens during weaning in infants aged 4-6 months could result in food tolerance, thus reducing the risk of developing allergies. OBJECTIVE: The aim of this study is to systematically review and carry out a meta-analysis of evidence on the outcome of early food introduction for preventing childhood allergic diseases. METHODS: We will conduct a systematic review of interventions through a comprehensive search of various databases including PubMed, Embase, Scopus, CENTRAL, PsycINFO, CINAHL, and Google Scholar to identify potential studies. The search will be performed for any eligible articles from the earliest published articles up to the latest available studies in 2023. We will include randomized controlled trials (RCTs), cluster RCTs, non-RCTs, and other observational studies that assess the effect of early food introduction to prevent childhood allergic diseases. RESULTS: Primary outcomes will include measures related to the effect of childhood allergic diseases (ie, asthma, allergic rhinitis, eczema, and food allergy). PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines will be followed for study selection. All data will be extracted using a standardized data extraction form and the quality of the studies will be assessed using the Cochrane Risk of Bias tool. A summary of findings table will be generated for the following outcomes: (1) total number of allergic diseases, (2) rate of sensitization, (3) total number of adverse events, (4) improvement of health-related quality of life, and (5) all-cause mortality. Descriptive and meta-analyses will be performed using a random-effects model in Review Manager (Cochrane). Heterogeneity among selected studies will be assessed using the I2 statistic and explored through meta-regression and subgroup analyses. Data collection is expected to start in June 2023. CONCLUSIONS: The results acquired from this study will contribute to the existing literature and harmonize recommendations for infant feeding with regard to the prevention of childhood allergic diseases. TRIAL REGISTRATION: PROSPERO CRD42021256776; https://tinyurl.com/4j272y8a. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): PRR1-10.2196/46816.

4.
J Clin Immunol ; 43(5): 999-1006, 2023 07.
Article in English | MEDLINE | ID: mdl-36882668

ABSTRACT

PURPOSE: Primary immunodeficiency disease (PID) affects various aspects of a patient's life. However, the health-related quality of life (HRQOL) of PID among Malaysian patients is poorly described. This study aimed to determine the quality of life of PID patients and their respective parents. METHOD: This cross-sectional study was performed from August 2020 to November 2020. Patients with PID and their families were invited to answer the PedsQL Malay version (4.0) questionnaire, the tool used to assess the HRQOL. A total of 41 families and 33 patients with PID answered the questionnaire. A comparison was performed with the previously published value of healthy Malaysian children. RESULT: Parents of respondents recorded a lower mean of total score than the parents of healthy children (67.26 ± 16.73 vs. 79.51 ± 11.90, p-value = 0.001, respectively). PID patients reported lower mean total score to healthy children (73.68 ± 16.38 vs. 79.51 ± 11.90, p-value = 0.04), including the psychosocial domain (71.67 ± 16.82 vs. 77.58 ± 12.63, p-value = 0.05) and school functioning (63.94 ± 20.87 vs. 80.00 ± 14.40, p-value = 0.007). No significant difference of reported HRQOL when comparing between subgroup of PID on immunoglobulin replacement therapy and those without immunoglobulin replacement (56.96 ± 23.58 vs. 65.83 ± 23.82, p-value 0.28). Socioeconomic status was found to be predictive of the lower total score of PedsQL in both parent and children reports. CONCLUSION: Parents and children with PID, especially those from middle socioeconomic status, have lower HRQOL and school function impairment than healthy children.


Subject(s)
Parents , Quality of Life , Child , Humans , Cross-Sectional Studies , Malaysia/epidemiology , Surveys and Questionnaires
5.
Mol Immunol ; 155: 44-57, 2023 03.
Article in English | MEDLINE | ID: mdl-36696839

ABSTRACT

INTRODUCTION: Goat's milk thought to be a good substitute for cow's milk protein allergic (CMPA) individuals. However, there is growing evidence that their proteins have cross-reactivities with cow's milk allergens. This study aimed to profile and compare milk proteins from different goat breeds that have cross-reactivity to cow's milk allergens. METHODOLOGY: Proteomics was used to compare protein extracts of skim milk from Saanen, Jamnapari, and Toggenburg. Cow's milk was used as a control. IgE-immunoblotting and mass spectrometry were used to compare and identify proteins that cross-reacted with serum IgE from CMPA patients (n = 10). RESULTS: The analysis of IgE-reactive proteins revealed that the protein spots identified with high confidence were proteins homologous to common cow's milk allergens such as α-S1-casein (αS1-CN), ß-casein (ß-CN), κ-casein (κ-CN), and beta-lactoglobulin (ß-LG). Jamnapari's milk proteins were found to cross-react with four major milk allergens: α-S1-CN, ß-CN, κ-CN, and ß-LG. Saanen goat's milk proteins, on the other hand, cross-reacted with two major milk allergens, α-S1-CN and ß-LG, whereas Toggenburg goat's milk proteins only react with one of the major milk allergens, κ-CN. CONCLUSION: These findings may help in the development of hypoallergenic goat milk through cross-breeding strategies of goat breeds with lower allergenic milk protein contents.


Subject(s)
Milk Hypersensitivity , Milk Proteins , Animals , Cattle , Female , Milk , Allergens , Goats , Proteomics , Immunoglobulin E , Caseins
6.
Front Immunol ; 14: 1308305, 2023.
Article in English | MEDLINE | ID: mdl-38283358

ABSTRACT

Background: Primary Immunodeficiency Disease (PID), also known as Inborn Errors of Immunity (IEI), comprises a group of rare genetic disorders that impair the body's immune responses. These conditions result from monogenic germline mutations that affect the function of genes governing the innate and adaptive immune system. Therefore, individuals with PID are more susceptible to infectious diseases, allergies, and autoimmune and autoinflammatory conditions. The prevalence of PID has been on the rise, with the number of classified diseases reaching 404, and 430 genetic defects reported to cause these conditions. However, in Malaysia, genetic testing for PID is currently limited and needs to be outsourced to overseas laboratories, posing financial challenges for families. Moreover, limited research has focused on the knowledge and awareness of genetic testing among parents of children with PID in Malaysia. This study aims to address this gap and provide valuable insights into the knowledge, awareness, and perception of genetic testing among this specific population. Method: This qualitative cross-sectional study utilised online open-ended, semi-structured focus group interviews to explore the perceptions and experiences of parents of children with Primary Immunodeficiency (PID). Participants were recruited through convenience sampling from the Malaysian Patient Organisation for Primary Immunodeficiencies (MyPOPI), a non-governmental organisation dedicated to providing support and raising awareness about PID. The study spanned from May 2023 to July 2023 and included participants from diverse regions of Malaysia who had undergone different diagnostic journeys in various hospitals. Result: The focus group discussions yielded 11 sub-themes that highlighted the experiences, understanding and challenges of the participants regarding genetic testing based on the semi-structured questions. These sub-themes were then grouped into four main themes that are awareness and understanding of genetic testing, the journey towards diagnosis and treatment, emotional impact and psychological factors, and the importance of medical experts in diagnosing and managing PID, as well as public perception and awareness. Conclusion: In conclusion, this study highlights the diverse knowledge, awareness, and perception surrounding genetic testing for PID. Factors such as access to services, family history, and personal circumstances shape individuals' understanding of genetic testing. The importance of healthcare professionals, along with the need for improved accessibility and targeted communication strategies, is underscored to enhance understanding and reduce stigma surrounding genetic testing for rare diseases like PID.


Subject(s)
Immunologic Deficiency Syndromes , Primary Immunodeficiency Diseases , Child , Humans , Immunologic Deficiency Syndromes/diagnosis , Immunologic Deficiency Syndromes/epidemiology , Immunologic Deficiency Syndromes/genetics , Malaysia/epidemiology , Cross-Sectional Studies , Genetic Testing , Parents , Rare Diseases/genetics , Primary Immunodeficiency Diseases/diagnosis , Primary Immunodeficiency Diseases/genetics , Perception
7.
Genes (Basel) ; 13(10)2022 10 19.
Article in English | MEDLINE | ID: mdl-36292785

ABSTRACT

Background: Inborn errors of immunity (IEIs) are comprised of heterogeneous groups of genetic disorders affecting immune function. In this report, a 17-month-old Malay patient suspected of having Hyper IgM syndrome, a type of IEIs, was described. However, the diagnosis of Hyper IgM syndrome was excluded by the normal functional studies and the mild features of ectodermal dysplasia observed from a further clinical phenotype inspection. Methods: Whole-exome sequencing (WES) was performed to unravel the causative mutation in this patient. Results: The variant analysis demonstrated a novel missense mutation in NFKBIA (NM_020529:c.94A > T,NP_065390:p.Ser32Cys) and was predicted as damaging by in silico prediction tools. The NFKBIA gene encodes for IκBα, a member of nuclear factor kappa B (NF-κB) inhibitors, playing an important role in regulating NF-κB activity. The mutation occurred at the six degrons (Asp31-Ser36) in IκBα which were evolutionarily conserved across several species. Prediction analysis suggested that the substitution of Ser32Cys may cause a loss of the phosphorylation site at residue 32 and a gain of the sumoylation site at residue 38, resulting in the alteration of post-translational modifications of IκBα required for NF-κB activation. Conclusion: Our analysis hints that the post-translational modification in the NFKBIA Ser32Cys mutant would alter the signaling pathway of NF-κB. Our findings support the usefulness of WES in diagnosing IEIs and suggest the role of post-translational modification of IκBα.


Subject(s)
Dysgammaglobulinemia , Ectodermal Dysplasia , Hyper-IgM Immunodeficiency Syndrome , Immunologic Deficiency Syndromes , Humans , NF-KappaB Inhibitor alpha/genetics , NF-kappa B/genetics , NF-kappa B/metabolism , Mutation, Missense , Ectodermal Dysplasia/genetics , I-kappa B Proteins/genetics , I-kappa B Proteins/metabolism
8.
J Asthma Allergy ; 15: 983-1003, 2022.
Article in English | MEDLINE | ID: mdl-35942430

ABSTRACT

The goal of allergic rhinitis (AR) management is to achieve satisfactory symptom control to ensure good quality of life. Most patients with AR are currently treated with pharmacotherapy. However, knowledge gaps on the use of pharmacotherapy still exist among physicians, particularly in the primary care setting, despite the availability of guideline recommendations. Furthermore, it is common for physicians in the secondary care setting to express uncertainty regarding the use of new combination therapies like intranasal corticosteroid plus antihistamine combinations. Inadequate treatment leads to significant reduction of quality of life that affects daily activities at home, work, and school. With these concerns in mind, a practical consensus statement was developed to complement existing guidelines on the rational use of pharmacotherapy in both the primary and secondary care settings.

9.
Front Immunol ; 13: 883446, 2022.
Article in English | MEDLINE | ID: mdl-35874699

ABSTRACT

To address inborn errors of immunity (IEI) which were underdiagnosed in resource-limited regions, our centre developed and offered free genetic testing for the most common IEI by Sanger sequencing (SS) since 2001. With the establishment of The Asian Primary Immunodeficiency (APID) Network in 2009, the awareness and definitive diagnosis of IEI were further improved with collaboration among centres caring for IEI patients from East and Southeast Asia. We also started to use whole exome sequencing (WES) for undiagnosed cases and further extended our collaboration with centres from South Asia and Africa. With the increased use of Next Generation Sequencing (NGS), we have shifted our diagnostic practice from SS to WES. However, SS was still one of the key diagnostic tools for IEI for the past two decades. Our centre has performed 2,024 IEI SS genetic tests, with in-house protocol designed specifically for 84 genes, in 1,376 patients with 744 identified to have disease-causing mutations (54.1%). The high diagnostic rate after just one round of targeted gene SS for each of the 5 common IEI (X-linked agammaglobulinemia (XLA) 77.4%, Wiskott-Aldrich syndrome (WAS) 69.2%, X-linked chronic granulomatous disease (XCGD) 59.5%, X-linked severe combined immunodeficiency (XSCID) 51.1%, and X-linked hyper-IgM syndrome (HIGM1) 58.1%) demonstrated targeted gene SS should remain the first-tier genetic test for the 5 common X-linked IEI.


Subject(s)
Agammaglobulinemia , X-Linked Combined Immunodeficiency Diseases , Agammaglobulinemia/diagnosis , Agammaglobulinemia/genetics , Child , Genetic Testing , High-Throughput Nucleotide Sequencing , Humans , Exome Sequencing , X-Linked Combined Immunodeficiency Diseases/genetics
11.
Front Pediatr ; 10: 846393, 2022.
Article in English | MEDLINE | ID: mdl-35433547

ABSTRACT

Introduction: Primary immunodeficiency diseases (PIDs) are chronic diseases that affect the various aspects of a patient's life. However, the impact of living with PIDs is poorly described. Objective: This study aimed to explore the living experience challenges among the Malaysian caregivers of the patients with PID who underwent a follow-up in the Universiti Sains Malaysia or those registered members of the Malaysian PIDs Society. Methodology: The study was conducted from March 1 to May 30, 2021. The parents of children with PIDs were invited to participate for a semi-structured in-depth interview at the PID clinics in the USM. The estimated time of each interview was 30 min. The semi-structured interview was performed via a telephone call because of COVID-19 pandemic restrictions. The audio recording of each interview was transcribed and translated from Malay to English. Subsequently, a thematic analysis utilizing the ATLAS.ti software was performed. Results: The thematic analysis revealed five main themes, which are living with fear and anxiety with four sub-themes (sickness, psychological issues, fear of infections and hereditary issues), PID healthcare support struggles with four sub-themes (PID health system, treatment, diagnosis and financial issues), knowledge with two sub-themes (educational issues and disease understanding), social constraint with two sub-themes (relationships and social isolations) and coping with three sub-themes (acceptance, child health improvement and emotional hygiene). Conclusion: Living with fear and anxiety is a major theme impacting the living experiences of Malaysian patients with PIDs. Improvements in healthcare delivery and disease education are needed to ensure optimal quality of life.

13.
Children (Basel) ; 8(11)2021 Nov 13.
Article in English | MEDLINE | ID: mdl-34828763

ABSTRACT

Food allergy has a significant impact on the quality of life (QoL) of children and can be measured using The Food Allergy Quality of Life Questionnaire-Parent Form (FAQLQ-PF). This study aimed to adapt, translate the FAQLQ-PF into Malay and determine the validity and reliability of the translated version. This cross-sectional questionnaire validation study was conducted among parents of children (0 to 12 years old) with food allergies across five sites in Selangor and Kuala Lumpur, Malaysia. The FAQLQ-PF-Malay underwent cross-cultural adaptation, translation, validation (content, face and construct) and reliability assessment. Exploratory factor analysis, internal consistency and test-retest reliability analyses were used to examine its construct validity and reliability. Out of 150 children, the majority were between the age of 7 to 12 years old (41%) and were female (81%). Three subscales were identified, which were: (i) social and dietary implication, (ii) food anxiety and (iii) emotional and physical impact. Four items were eliminated because of weak factor loadings. The Cronbach's alpha for each subscale ranged from 0.88 to 0.94, with an overall Cronbach's alpha of 0.95. The intra-class correlation coefficient ranged from 0.54 (95% CI: 0.10-0.77) to 0.97 (95% CI: 0.90-0.99). The 26-item FAQLQ-PF-Malay retained the three-factor structure of the original FAQLQ-PF. The FAQLQ-PF-Malay is a valid and reliable tool to assess the QoL of Malaysian children with food allergies.

14.
Allergy Asthma Clin Immunol ; 17(1): 50, 2021 May 17.
Article in English | MEDLINE | ID: mdl-34001231

ABSTRACT

BACKGROUND: A retrospective review of clinical manifestations and demographic pattern of patients diagnosed as chronic granulomatous disease (CGD) from 7 hospitals in Malaysia. An analysis of the available database would establish clinical characteristics, diagnoses and outcome including microbiologic pattern. Studying the demography allows us to document the occurrence of CGD amongst multiethnic groups and its geographical distribution for Malaysia. METHODS: Data from the Malaysia Primary Immunodeficiency Network (MyPIN) with cases of CGD diagnosed from 1991 until 2016 were collated and analysed. RESULTS: Twenty patients were diagnosed as CGD. Males (N = 13, 65%) outnumber females (N = 7, 35%). CGD is commonest amongst the Malays (65%) followed by the Chinese (15.0%), Indians (10.0%) and natives of Borneo (10.0%), reflecting the ethnic composition of the country. The mean age of diagnosis was 3.7 years. There was a positive family history in 40% of the cases. Abscess was the main presenting feature in 16 patients (80%) with one involving the brain. Pneumonia occurred in 10 (50%) and one with complicated bronchiectasis. Catalase-positive bacteria were the most commonly isolated pathogen with Chromobacterium violaceum predominating (N = 5, 25%) with consequent high mortality (N = 4, 80%). All CGD patients with C. violaceum infection displayed CD4 + (T helper cells) lymphopenia. CONCLUSION: This study has shown CGD occurs in the major ethnic groups of Malaysia. To the best of our knowledge, this is the first and the largest series of chronic granulomatous disease in South East Asia which may be reflective of similar clinical pattern in the region. C. violaceum infection is associated with a higher mortality in CGD patients in Malaysia. All the CGD patients with C. violaceum infection in this patient series displayed CD4 + (T helper) lymphopenia. We recorded rare clinical manifestation of CGD viz. brain abscess and bronchiectasis.

15.
J Nutr Sci ; 10: e108, 2021.
Article in English | MEDLINE | ID: mdl-35059189

ABSTRACT

Children with cerebral palsy (CP) typically suffer from congenital deformities, such as scoliosis and contractures, therefore, it is a challenge to measure the stature of CP children. Studies have suggested that predictive equations based on tibia length (TL) may be used as an alternative method in measuring the actual height or stature. The present study aimed to develop and validate predictive equations based on TL for CP children in Malaysia across all five levels of gross motor functions (GMFCS I to V) through a cross-sectional study. All subjects were recruited from Hospitals and Community-Based Rehabilitation (CBR) in the central and southern regions of Malaysia. Two predictive equation models were developed using multiple linear regression. For Model 1, the predictive equation was developed based on TL. On the other hand, Model 2 was developed based on TL with age was included. A flexible Seca measuring tape was used to measure the stature and TL. CP children aged 2-18 years were classified into the equation development group (EDG), n 177 and the validation group (VG), n 139. Model 1, Height = 32⋅3 + 3⋅14 (TL), demonstrated a strong correlation with the actual height (R2 0⋅834), small SEE (1⋅42), and high intra correlation coefficient (0⋅929). The findings suggested that Model 1 was more accurate in estimating the height of CP children aged 2-18 years. This model was shown to suit the Malaysian population and applicable across all GMFCS levels.


Subject(s)
Cerebral Palsy , Adolescent , Body Height , Child , Child, Preschool , Cross-Sectional Studies , Humans
16.
Br J Clin Pharmacol ; 87(7): 2956-2966, 2021 07.
Article in English | MEDLINE | ID: mdl-33377197

ABSTRACT

AIMS: There is considerable interpatient variability in the pharmacokinetics (PK) of intravenous immunoglobulin G (IVIG), causing difficulty in optimizing individual dosage regimen. This study aims to estimate the population PK parameters of IVIG and to investigate the impact of genetic polymorphism of the FcRn gene and clinical variability on the PK of IVIG in patients with predominantly antibody deficiencies. METHODS: Patients were recruited from four hospitals. Clinical data were recorded and blood samples were taken for PK and genetic studies. Population PK parameters were estimated by nonlinear mixed-effects modelling in Monolix®. Models were evaluated using the difference in objective function value, goodness-of-fit plots, visual predictive check and bootstrap analysis. Monte Carlo simulation was conducted to evaluate different dosing regimens for IVIG. RESULTS: A total of 30 blood samples were analysed from 10 patients. The immunoglobulin G concentration data were best described by a one-compartment model with linear elimination. The final model included both volume of distribution (Vd) and clearance (CL) based on patient's individual weight. Goodness-of-fit plots indicated that the model fit the data adequately, with minor model mis-specification. Genetic polymorphism of the FcRn gene and the presence of bronchiectasis did not affect the PK of IVIG. Simulation showed that 3-4-weekly dosing intervals were sufficient to maintain IgG levels of 5 g L-1 , with more frequent intervals needed to achieve higher trough levels. CONCLUSIONS: Body weight significantly affects the PK parameters of IVIG. Genetic and other clinical factors investigated did not affect the disposition of IVIG.


Subject(s)
Immunoglobulins, Intravenous , Models, Biological , Administration, Intravenous , Computer Simulation , Humans , Immunoglobulins, Intravenous/pharmacokinetics , Monte Carlo Method
17.
Article in English | MEDLINE | ID: mdl-32971369

ABSTRACT

The main challenges in the purification of αS2-casein are due to the low quantity in milk and high homology with other casein subunits, i.e., αS1-casein, ß-casein, and κ-casein. To overcome these challenges, the aim of this study was to develop a two-step purification to isolate native αS2-casein in goat milk from five different breeds; British Alpine, Jamnapari, Saanen, Shami, and Toggenburg. The first step of the purification was executed by anion-exchange chromatography under optimal elution conditions followed by size exclusion chromatography. Tryptic peptides from in-gel digestion of purified αS2-casein were sequenced and analyzed by LC-ESI-MS/MS. From 1.05 g of whole casein, the highest yield of αS2-casein (6.7 mg/mL) was obtained from Jamnapari and the lowest yield (2.2 mg/mL) was from Saanen. A single band of pure αS2-casein was observed on SDS-PAGE for all breeds. The αS2-casein showed coverage percentage of amino acid sequence from 76.68 to 92.83%. The two-step purification process developed herein was successfully applied for isolating native αS2-casein from goat milk with high purity, which will allow for future in vitro studies to be conducted on this protein.


Subject(s)
Caseins , Chromatography, Liquid/methods , Goats/classification , Milk/chemistry , Animals , Caseins/analysis , Caseins/chemistry , Caseins/classification , Caseins/isolation & purification , Goats/metabolism , Milk/classification , Spectrometry, Mass, Electrospray Ionization/methods , Tandem Mass Spectrometry/methods
19.
Nutrients ; 12(8)2020 Aug 12.
Article in English | MEDLINE | ID: mdl-32806653

ABSTRACT

Allergic diseases are the most common chronic illness in childhood. Findings from developed countries have reported associations between Vitamin D levels during pregnancy and offspring allergy risk. This prospective cohort study aimed to determine the associations between maternal Vitamin D levels during late pregnancy and allergic diseases in Malaysian infants during the first year of life. Serum 25(OH)D concentrations of 380 pregnant women in the third trimester were measured using a chemiluminescent immunoassay. Children's allergic outcomes were assessed at 3, 6, and 12 months based on parental reports. Specific IgE antibodies against food and inhalant allergens were measured in infants at 12 months of age. A total of 43.2% pregnant women were Vitamin D deficient (<30 nmol/L) and 56.8% were nondeficient (≥30 nmol/L). A total of 27.6% of the infants had eczema, 6.1% had wheeze, 27.4% had food sensitization, 10.8% had inhalant allergen sensitization, and 3.8% had IgE-mediated food allergy during the first year of life. Compared with the nondeficient group, maternal Vitamin D deficiency in late pregnancy was not associated with any allergic outcomes after adjustment for potential confounding factors. In conclusion, the present study does not support an association between maternal Vitamin D levels in late pregnancy and allergic outcomes during the first year of life.


Subject(s)
Hypersensitivity/etiology , Pregnancy Complications/blood , Prenatal Exposure Delayed Effects/etiology , Vitamin D Deficiency/complications , Vitamin D/analogs & derivatives , Adult , Autoantibodies/blood , Female , Humans , Hypersensitivity/epidemiology , Immunoglobulin E/immunology , Infant , Malaysia/epidemiology , Male , Pregnancy , Prenatal Exposure Delayed Effects/epidemiology , Prevalence , Prospective Studies , Vitamin D/blood
20.
Molecules ; 25(11)2020 Jun 05.
Article in English | MEDLINE | ID: mdl-32516919

ABSTRACT

The chemical, technological and allergy properties of goat's milk are significantly affected by the level of αs1-casein. Detection and quantification of αs1-casein requires high-specificity methods to overcome high-sequence similarity between this protein and others in the casein family. Unavailability of antibodies with high affinity and specificity towards goat αs1-casein hinders the development of immuno-based analytical methods such as enzyme-linked immunosorbent assay (ELISA) and biosensors. Here, we report the generation of polyclonal antibodies (or immunoglobulins, IgGs) raised towards goat αs1-casein N- (Nter) and C-terminal (Cter) peptide sequences. The Nter and Cter peptides of goat αs1-casein were immunized in rabbits for the generation of antisera, which were purified using protein G affinity chromatography. The binding affinity of the antisera and purified IgGs were tested and compared using indirect ELISA, where peptide-BSA conjugates and goat αs1-casein were used as the coating antigens. The Nter antiserum displayed higher titer than Cter antiserum, at 1/64,000 and 1/32,000 dilutions, respectively. The purification step further yielded 0.5 mg/mL of purified IgGs from 3 mL of antisera. The purified Nter IgG showed a significantly (p < 0.05) higher binding affinity towards peptide-BSA and goat αs1-casein, with lower Kd value at 5.063 × 10-3 µM compared to 9.046 × 10-3 µM for the Cter IgG. A cross-reactivity test showed that there was no binding in neither Nter nor Cter IgGs towards protein extracts from the milk of cow, buffalo, horse and camel. High-quality antibodies generated will allow further development of immuno-based analytical methods and future in vitro studies to be conducted on goat αs1-casein.


Subject(s)
Antibodies/immunology , Antibodies/metabolism , Caseins/analysis , Caseins/immunology , Milk/chemistry , Peptide Fragments/immunology , Animals , Antibodies/isolation & purification , Enzyme-Linked Immunosorbent Assay , Female , Goats , Immunization , Rabbits
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